Expression and function of cyclooxygenase and lipoxygenase enzymes in human islets of Langerhans.

Arachidonic acid (AA) is generated in pancreatic beta-cells through the activation of Ca2+-dependent cytosolic phospholipase A2 (cPLA2) and the consequent hydrolysis of membrane phospholipids in the sn-2 position of the glycerophospholipid backbone. AA acts as a second messenger in beta-cells to elevate cytosolic Ca2+ levels and stimulate insulin secretion, but it is not clear whether these are direct effects of AA or are dependent on its metabolism by cyclooxygenase (COX) and/or lipoxygenase (LOX) enzymes. In addition, much of the published data in this area have been generated using insulin-secreting cell lines or rodent islets, with very little information on AA generation and metabolism in human islets of Langerhans.

The role of arachidonic acid and its metabolites in insulin secretion from human islets of langerhans.

The roles played by arachidonic acid and its cyclooxygenase (COX)-generated and lipoxygenase (LOX)-generated metabolites have been studied using rodent islets and insulin-secreting cell lines, but very little is known about COX and LOX isoform expression and the effects of modulation of arachidonic acid generation and metabolism in human islets. We have used RT-PCR to identify mRNAs for cytosolic phospholipase A(2) (cPLA(2)), COX-1, COX-2, 5-LOX, and 12-LOX in isolated human islets. COX-3 and 15-LOX were not expressed by human islets.

Glucose-induced regulation of COX-2 expression in human islets of Langerhans.

Cyclo-oxygenase (COX), the enzyme responsible for conversion of arachidonic acid to prostanoids, exists as two isoforms. In most tissues, COX-1 is a constitutive enzyme involved in prostaglandin-mediated physiological processes, whereas COX-2 is thought to be induced by inflammatory stimuli. However, it has previously been reported that COX-2 is the dominant isoform in islets and an insulin-secreting beta-cell line under basal conditions.

The role of cytosolic phospholipase A(2) in insulin secretion.

Cytosolic phospholipase A(2) (cPLA(2)) comprises a widely expressed family of enzymes, some members of which have the properties required of signal transduction elements in electrically excitable cells. Thus, alpha- and beta-isoforms of cPLA(2) are activated by the increases in intracellular Ca(2+) concentration ([Ca(2+)](i)) achieved in depolarized cells. Activation is associated with a redistribution of the enzyme within the cell; activation of cPLA(2) generates arachidonic acid (AA), a biologically active unsaturated fatty acid that can be further metabolized to generate a plethora of biologically active molecules.

A key role for beta-cell cytosolic phospholipase A(2) in the maintenance of insulin stores but not in the initiation of insulin secretion.

Cytosolic phospholipase A(2) (cPLA(2)) is a Ca(2+)-sensitive enzyme that has been implicated in insulin secretion in response to agents that elevate beta-cell intracellular Ca(2+) ([Ca(2+)](i)). We generated clones of the MIN6 beta-cell line that stably underexpress cPLA(2) by transfection with a vector in which cPLA(2) cDNA had been inserted in the antisense orientation. Reduced expression of cPLA(2) was confirmed by Western blotting.